Scammell Project Summary In patients with obstructive sleep apnea (OSA), arousals play a life-saving role, and we hypothesize that neurons in the laterodorsal and pedunculopontine tegmental nuclei (LDT/PPT) play an essential role in many aspects of arousal, including arousals from sleep in OSA. Our goals are to determine the specific functions of the LDT/PPT neurons producing acetylcholine, glutamate and GABA; to map the anatomic projections of these cells; and to examine how they influence the activity of key target regions that regulate sleep/wake behavior and respiration. We will pursue these goals using optogenetics to examine acute, brief increases or decreases in neuronal activity within a behavioral state, and pharmacosynthetics using Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) to examine changes lasting hours. As a model of OSA, many experiments will use the Repetitive CO2 Arousal (RCA) method to examine responses to hypercapnia during sleep. To determine if specific LDT/PPT neurons are sufficient for wakefulness, EEG activation, and the response to RCA, we will activate each class of neurons using the hM3 DREADD or channelrhodopsin (ChR2). To determine if specific LDT/PPT neurons are necessary for wakefulness, EEG activation, and the response to RCA, we will inhibit each class of neurons using the hM4 DREADD or ArchT, an inhibitory archaerhodopsin. To define the key LDT/PPT target regions mediating these behavioral responses, we will map projections of specific LDT/PPT neurons using conditional anterograde and retrograde tracing; determine which projections form functional synapses using ChR2- Assisted Circuit Mapping (CRACM); and determine which target regions are functionally most important by focally stimulating or inhibiting glutamatergic and cholinergic nerve terminals in each target region using ChR2 or ArchT. Collectively, these multidisciplinary experiments will define the neural mechanisms through which specific types of LDT/PPT neurons promote wakefulness, cortical activation, and arousals in a mouse model of OSA. The results of these experiments should substantially improve our scientific knowledge of the neural mechanisms that generate wakefulness and arousals due to hypercapnia, and ultimately lead to better treatments for obstructive sleep apnea.